Functional assays with CRISPR-Cas9 knockout for the CG9068 gene in Drosophila melanogaster: a model for subfunctionalization and sex conflict

• Grant number: 24/05885-5
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor
• Start date: November 01, 2024
• End date: October 31, 2025
• Field of knowledge: Biological Sciences - Genetics - Animal Genetics
• Principal Investigator: Rodrigo Cogni
• Grantee: Eduardo Guimarães Dupim
• Supervisor: Manyuan Long
• Host Institution: Instituto de Biociências (IB). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Institution abroad: University of Chicago, United States
• Associated to the scholarship: 20/04880-9 - Evolutionary convergence in duplicative transposition and subfunctionalization of the male fertility gene kl-2 in drosophilids, BP.PD

Abstract

Subfunctionalization is considered one of the most frequent models for maintaining gene duplications, remaining an important source of evolutionary novelties through the emergence of new genes. In this model, each copy specializes in part of the original gene's functions. Also, this process might be a resolution of sexual conflict (SC), when a gene has conflicting roles between isoforms and/or expression in different sexes. Though widely predicted, there have been few reported cases of strict subfunctionalization and sexual conflict with solid experimental evidence. In this internship project, we propose a functional study of the CG9068 gene, a subfunctionalized gene from Drosophila fruit flies, potentially involved with SC. CG9068 is the autosomal copy of the kl-2, a Y chromosome gene that codifies a dynein protein involved with spermatozoa ciliary movement. kl-2 was ancestrally located in an autosome before being transposed by DNA-based duplication to the Y chromosome in the Drosophila ancestral, leaving an autosomal copy (CG9068) in its ancestral position. CG9068 has been reduced in size, encoding only the first domain of the original protein. Recently, our analyses in D. melanogaster show CG9068 is expressed in auditory cells in the antenna - indicating that the shortened dynein protein codified by this gene could be involved with the ciliary motility of these cells, fundamental for the hearing and proprioception senses of both sexes. This function justifies the maintenance and subfunctionalization of CG9068 after the duplicative transposition of kl-2 to the Y chromosome, where the gene specialized in sperm functions. To confirm this hypothesis, we will investigate CG9068 functions, through CRISPR-Cas9 knockout lines and functional assays for hearing and fertility. Also, in the SC scenario, vestigial expression of CG9068 in sperm could lower males' fitness, and males without the CG9068 gene could increase their fitness, explaining the transposition to Y as a solution for this sexual conflict.